This invention relates generally to the synthesis of chemical compounds, and more particularly, to the solid phase synthesis of combinatorial libraries of chemical compounds.
Combinatorial organic synthesis is becoming an important tool in drug discovery. Methods for the synthesis of large numbers of diverse compounds have been described (Ellman et al., Chem. Rev. 96, 555-600 (1996)), as have methods for tagging systems (Ohlmeyer et al., Proc. Natl. Acad. Sci. USA 90, 10922-10926 (1993)). The growing importance of combinatorial synthesis has created a need for new resins and linkers having chemical and physical properties that accommodate a wide range of conditions. Success in combinatorial synthesis on solid phase supports depends on the ability to synthesize diverse sets of molecules and to then cleave those molecules from the supports cleanly and in good yield.
Linkers are molecules that are attached to a solid support and to which the desired members of a library of chemical compounds may in turn be attached. When the construction of the library is complete, the linker allows clean separation of the target compounds from the solid support without harm to the compounds and preferably without damage to the support. Several linkers have been described in the literature. Their value is constrained by the need to have sufficient stability which allows the steps of combinatorial synthesis under conditions that will not cleave the linker. An additional constraint is the need to have a fairly high lability under at least one set of conditions that is not employed in the chemical synthesis.
For example, if an acid labile linker is employed, then the combinatorial synthesis must be restricted to reactions that do not require the presence of an acid of sufficient strength to endanger the integrity of the linker. Likewise, when a photocleavable linker is employed, conditions that exclude light are necessary to avoid untimely cleavage of the compound from the resin. This sort of balancing act often imposes serious constraints on the reactions that are chosen for preparation of the library.
The 4-[4-(hydroxymethyl)-3-methoxyphenoxy]butyryl residue is a known linker, which is attached to a solid support having amino groups by forming an amide with the carboxyl of the butyric acid chain. N-Protected amino acids are attached to the hydroxyl of the 4-hydroxymethyl group via their carboxyl to form 2,4-dialkoxybenzyl esters, which can be readily cleaved in acid media when the synthesis is complete (Riniker et al., Tetrahedron 49, 9307-9312 (1993)). The drawback to such 2,4-dialkoxybenzyl esters is their instability with many of the reagents that are available for use in combinatorial synthesis resulting in cleavage of the ester.
A somewhat more stable ester is formed from 4-[4-(hydroxymethyl)phenoxy]butyric acid, described in European published application EP 445915. In this case, the ester was cleaved with a 90:5:5 mixture of trifluoroacetic acid, dimethyl sulfide and thioanisole.
When the desired product is a peptide amide, the 4-[4-(formyl)-3,5-dimethoxyphenoxy]butyryl residue has been employed as a linker. This particular linker is attached to a solid phase substrate via the carboxyl of the butyric acid chain, and the 4-formyl group is reductively aminated. N-Protected amino acids are then reacted with the alkylamine via their carboxyl to form 2,4,6-trialkoxybenzylamides. These may be cleaved by 1:1 trifluoroacetic acid in dichloromethane (PCT application WO97/23508).
If a photocleavable linker is used to attach chemical compounds to the main support, milder photolytic conditions of cleavage can be used which complement traditional acidic or basic cleavage techniques. A wider range of combinatorial synthetic conditions will be tolerated by photocleavable linkers (Gallop et al., J. Med. Chem. 37, 1233-1251 (1994); Gordon et al., J. Med. Chem. 37, 1385-1401 (1994)).
A phenacyl based linking group that is photocleavable has been described (Wang et al., J. Org. Chem. 41, 3258 (1976)). The 4-bromomethyl-3-nitrobenzoyl residue has been widely employed as a photocleavable linker for both peptide acids and amides (Rich et al., J. Am. Chem. Soc. 97, 1575-1579 (1975); Hammer et al., Int. J. Peptide Protein Res. 36, 31-45 (1990)). This linker suffers from unduly slow cleavage rates, with typical photolysis times in organic solvents ranging from 12 to 24 hours. Moreover, photolytic cleavage of the linker generates a reactive and chromogenic nitrosoaldehyde on the resin support which can trap liberated compounds (Patchnornik et al., J. Am. Chem. Soc. 92, 6333-6335 (1970)). An xcex1-methyl-2-nitrobenzyl linker was designed to obviate formation of the nitroso-aldehyde, but inefficient release of pentapeptides resulted due to swelling of the resin support (Ajajaghosh et al., Tetrahedron 44, 6661-6666 (1988)). Photocleavable linkers such as the 3-bromomethyl-4-nitro-6-methoxyphenoxyacetyl residue are stable to acidic or basic conditions yet, are rapidly cleavable under mild conditions and do not generate highly reactive byproducts (U.S. Pat. No. 5,739,386, issued Apr. 14, 1998).
It would be useful to have a linker-resin combination that would withstand a wider range of reaction conditions in combinatorial synthesis, but that could be readily and cleanly cleaved following completion of the solid phase synthesis.
The present invention relates to a linker-resin combination that demonstrates the ability to withstand many common reaction conditions and yet is cleavable under relatively mild conditions. In the following disclosure, the variables are defined when introduced and retain that definition throughout.
In one aspect, the invention relates to a substrate for solid phase synthesis comprising a solid phase-linker combination of the formula I: 
wherein: 
represents the residue of a solid support having a plurality of amino groups and the remainder constitutes the linker;
R1 is xe2x80x94NO2 or xe2x80x94CHO;
R2 is xe2x80x94OCH3, xe2x80x94CHO or xe2x80x94H; and
n=1 or 3-12.
Preferred solid phase supports are aminomethylated poly(styrene-co-divinylbenzene) and divinylbenzene-cross-linked resin, polyethyleneglycol-grafted polystyrene functionalized with amino groups.
In another aspect, the invention relates to a chemical intermediate of the formula II: 
wherein:
R1 is xe2x80x94NO2 or xe2x80x94CHO;
R2 is xe2x80x94OCH3, xe2x80x94CHO or xe2x80x94H;
R3 is chosen from the group consisting of hydroxyl, the residue of a solid support having a plurality of amino groups, and the residue of an ester; and n=1 or 3-12. A preferred ester residue is t-butoxy.
In a further aspect, the invention relates to processes for preparing the foregoing substrate for solid phase synthesis. One process comprises combining in a suitable solvent, a coupling agent, a solid support having a plurality of amino groups, and a compound of formula II: 
wherein R3 is hydroxyl. A preferred process is combining in a suitable solvent, a coupling agent, a solid support having a plurality of amino groups, and a compound of formula II wherein R1 is xe2x80x94NO2, R2 is xe2x80x94CHO and R3 is hydroxyl. In another preferred process, R1 is xe2x80x94CHO and R2 is xe2x80x94OCH3. In yet another preferred process, R1 is xe2x80x94CHO and R2 is xe2x80x94H.
In yet another aspect, the invention relates to a process for solid phase synthesis comprising:
a) reacting a substrate for solid phase synthesis of the formula I: 
wherein: 
represents the residue of a solid support having a plurality of amino groups and the remainder constitutes the linker;
R1 is xe2x80x94NO2 or xe2x80x94CHO;
R2 is xe2x80x94OCH3, xe2x80x94CHO or xe2x80x94H; and
n=1 or 3-12,
with a reagent capable of reacting with an aldehyde to provide a support-linked synthon;
b) carrying out a plurality of chemical transformations on said support-linked synthon to provide a support-linked product; and
c) treating said support-linked product with a condition of ultraviolet light or acid to cleave the product from the support and linker. When R1xe2x95x90xe2x80x94NO2 and R2xe2x95x90xe2x80x94CHO, the support-linked product is cleaved with ultraviolet light. When R1xe2x95x90xe2x80x94CHO and R2xe2x95x90xe2x80x94OCH3, the support-linked product is cleaved by mild acid. When R1xe2x95x90xe2x80x94CHO and R2xe2x95x90xe2x80x94H, the support-linked product is cleaved by a stronger concentration of acid. Trifluoroacetic acid is a preferred acid for cleavage.
In another aspect, the invention relates to a process for preparing a substrate for solid phase synthesis comprising:
combining in a suitable solvent, a coupling reagent, a solid support having a plurality of amino groups, a compound of the formula: 
and a compound of the formula: 
to produce said substrate of formula III: 
wherein: 
represents the residue of a solid support having a plurality of amino groups and the remainder constitutes the linker; R2xe2x95x90xe2x80x94OCH3 or xe2x80x94H; and n=1 or 3-12.
In yet a further aspect, the invention relates to a process for solid phase synthesis comprising:
a) reacting a substrate for solid phase synthesis of the formula III: 
wherein: 
represents the residue of a solid support having a plurality of amino groups and the remainder constitutes a linker; R2xe2x95x90xe2x80x94OCH3 or xe2x80x94H; and n=1 or 3-12, with a reagent capable of reacting with an aldehyde to provide a support-linked synthon;
b) carrying out a plurality of chemical transformations on said support-linked synthon to provide a support-linked product;
c) treating said support-linked product with ultraviolet light to cleave the photocleavable support-linked product from the support and linker; and
d) treating said support-linked product with trifluoroacetic acid to cleave the acid cleavable support-linked product from the support and linker.
When R2xe2x80x94OCH3, trifluoroacetic acid (2-25%) in CH2Cl2 cleaves the support-linked product. When R2xe2x95x90xe2x80x94H, trifluoroacetic acid (50-100%) in CH2Cl2 cleaves the support-linked product.